Objective—To determine the anisotropic characteristics of the microarchitecture of the subchondral bone (SCB) plate and trabecular bone (TBB) of the distopalmar aspect of the metacarpal condyles in horses with different stages of SCB disease.
Sample Population—12 third metacarpal bone pairs from racing Thoroughbreds euthanized for diverse reasons.
Procedures—Both metacarpi were collected from horses with SCB changes that were mild (sclerosis and focal radiolucencies; n = 6) or severe (multifocal radiolucencies and articular surface defects; 6). Sample blocks of SCB plate and TBB were collected from the distopalmar aspect of both condyles and the sagittal ridge and examined via 3-D micro-computed tomography at 45-?m isotropic voxel resolution. For each sample, the angle between the principal orientation of trabeculae and the sagittal plane and the degree of anisotropy (DA) were calculated from mean intercept length measurements.
Results—Condylar samples had significantly lower angle (mean, 8.9°; range, 73° to 10.9°) than sagittal ridge samples (mean, 40.7°; range, 33.6° to 49.2°), TBB had significantly higher DA (mean ± SE, 1.75 ± 0.04) than SCB plate (1.29 ± 0.04), and mildly diseased TBB had higher DA (1.85 ± 0.06) than severely diseased TBB (1.65 ± 0.06).
Conclusions and Clinical Relevance—The highly ordered appearance of trabeculae within the condyles supports the concept that joint loading is primarily transmitted through the condyles and not the sagittal ridge. The sharp changes in the trajectories of the SCB trabeculae at the condylar grooves may be indicative of hypothetical tensile forces at this location contributing to the pathogenesis of condylar fractures. (Am J Vet Res 2010;71:1148—1153)
Objective—To evaluate the effects of peroxisome proliferator-activated receptor gamma (PPARγ) agonists on tissue metabolism in cartilage and synovial explants from dogs.
Sample Population—Cartilage-synovial membrane expiants from 12 dogs.
Procedures—Expiants were cultured for 21 days without (negative control) or with interleukin (IL)-1β (positive control) or with IL-1β and 2 concentrations of 2 PPARγ agonists (15-deoxy-Δ 12,14-prostaglandin J2 [PGJ2] and pioglitazone). Media were collected on days 3, 7, 9, 12, 15, 18, and 21 and assessed for glycosaminoglycan (GAG), nitric oxide (NO), and prostaglandin E2 (PGE2) concentrations. Tissue GAG and hydroxyproline concentrations were determined in cartilage expiants collected on day 21.
Results—The GAG concentrations of cartilage expiants cultured in IL-1β (100 ng/mL) with 2 concentrations of PGJ2 were significantly higher than those in all other groups, whereas media GAG concentrations were significantly lower in the high-concentration PGJ2-treated groups, compared with all other groups. The PGE2 concentrations were significantly lower in the PGJ2 treatment groups, compared with the positive control and the pioglitazone treatment groups on days 3 to 21. The NO concentrations were significantly lower in PGJ2 treatment groups, compared with the other groups on days 3 and 12 to 21.
Conclusions and Clinical Relevance—PGJ2, an endogenous PPARγ agonist, may have anti-inflammatory and chondroprotective effects in an osteosteoarthritic joint environment (Am J Vet Res 2010;71:1142-1147)
Objective—To evaluate chondrocyte death in canine articular cartilage exposed in vitro to bupivacaine with and without methylparaben and to compare viability for cartilage with intact or mechanically debrided surfaces.
Sample Population—Both glenohumeral joints from 10 adult canine cadavers.
Procedures—10 osteochondral cores were harvested from each of the 20 humeral heads; synovium and 1 core from each joint were examined to verify joint health, and the other 9 cores were exposed to canine chondrocyte culture medium (CCCM), a 0.5% solution of bupivacaine, or 0.5% solution of bupivacaine with methylparaben for 5, 15, or 30 minutes.
Results—For the superficial zone of surface-intact chondrocytes, bupivacaine with methylparaben caused a significantly higher percentage of chondrocyte death at 5 minutes (47.7%) than did bupivacaine (23.6%) or CCCM (25.4%). Bupivacaine (53.8%) and bupivacaine with methylparaben (62.5%) caused a significantly higher percentage of chondrocyte death at 30 minutes than did CCCM (20.0%). For the superficial zone of chondrocytes with debrided surfaces, bupivacaine with methylparaben caused a significantly higher percentage of chondrocyte death at 30 minutes (59%) than it did at 5 minutes (37.7%). Bupivacaine with methylparaben caused a significantly higher percentage of chondrocyte death at 30 minutes (59.0%) than did CCCM (28.9%). For middle and deep zones of chondrocytes, treatment solution and surface debridement had minimal effects on percentage of chondrocyte death.
Conclusions and Clinical Relevance—Bupivacaine and bupivacaine with methylparaben were cytotoxic to canine articular chondrocytes in vitro. Intra-articular administration of bupivacaine is not recommended for clinical use until additional studies are conducted.
Objective—To study radiographic and genetic aspects of hereditary radial head subluxation in Bouviers des Flandres.
Animals—26 related Bouviers des Flandres affected with bilateral subluxation of the radial head, 10 unaffected related dogs, and 29 unrelated Bouviers des Flandres with diagnoses of nonskeletal diseases.
Procedures—All dogs were radiographically studied, and their DNA was analyzed with a genome-wide screen of 1,536 single nucleotide polymorphisms. In addition, karyotyping was performed in an unaffected dam and its affected offspring.
Results—Both forelimbs of affected dogs were disproportionately short with caudolateral subluxation or luxation of the radial head. Angulation of the radial axis at the mid-diaphysis ranged from 9.3° to 30.3° (mean ± SD, 14.9 ± 6.1°), with an estimated age of onset from 0 to 4 months. Poorly defined medial coronoid processes and osteoarthritis of the elbow joint, cranial bowing of the olecranon, and disturbed growth in length of the ulna with sharply demarcated spurs were noticed on radiographs of affected dogs. Genealogical analysis indicated that most affected dogs were closely related, but the mode of inheritance was not clear. The DNA analysis found that 205 single nucleotide polymorphisms were monomorphic in the affected dogs. Conventional chromosome staining revealed no numerical chromosomal aberration.
Conclusions and Clinical Relevance—Congenital radial head luxation and subluxation in the studied Bouviers des Flandres were characterized by angulation of the radial axis leading to caudolateral subluxation of the radial head and insufficient growth of the distal portion of the ulna together with cranial bowing of the olecranon.
Objective—To determine the effects of horse age, osteochondral injury, and joint type on a synthesis biomarker and 3 degradative biomarkers of type II collagen in Thoroughbreds.
Animals—Healthy rested adult (3- to 12-year-old) Thoroughbreds (n = 19), yearling (1- to 2-year-old) Thoroughbreds (40), and Thoroughbred racehorses (2 to 7 years old) undergoing arthroscopic surgery for removal of osteochondral fragments that resulted from training or racing (41).
Procedures—Samples of blood and metacarpophalangeal, metatarsophalangeal, or carpal joint synovial fluid (SF) were collected from all horses. Commercially available assays were used to analyze SF and serum concentrations of type II collagen biomarkers of synthesis (carboxy propeptide of type II collagen [CPII]) and degradation (cross-linked C-telopeptide fragments of type II collagen [CTX II], neoepitope generated by collagenase cleavage of type I and II collagen [C1,2C], and neoepitope generated by collagenase cleavage of type II collagen [C2C]).
Results—Osteochondral injury affected concentrations of CPII, CTX II, C1,2C, and C2C in SF, serum, or both, compared with concentrations in healthy adult horses. Compared with adult horses, yearling horses had increased SF or serum concentrations of degradative biomarkers (CTX II, C1,2C, and C2C). Concentrations were higher in carpal than metacarpophalangeal or metatarsophalangeal joints for all biomarkers in osteochondral-injured horses. Variable differences in SF concentrations between joint types were detected in healthy adult and yearling horses.
Conclusions and Clinical Relevance—Horse age, osteochondral injury, and joint type all significantly affected type II collagen biomarker concentrations in SF and serum of Thoroughbreds.
Objective—To compare macrostructural and microstructural features of proximal sesamoid bones (PSBs) from horses with and without PSB midbody fracture to gain insight into the pathogenesis of PSB fracture.
Sample Population—PSBs from 16 Thoroughbred racehorses (8 with and 8 without a PSB midbody fracture).
Procedures—Parasagittal sections of fractured and contralateral intact PSBs from horses with a PSB fracture and an intact PSB from age- and sex-matched control horses without a PSB fracture were evaluated for visual, radiographic, microradiographic, histologic, and his-tomorphometric differences in bone porosity, vascular channels, heme pigment, trabecular anisotropy, and pathological findings.
Results—Fractured PSBs and their contralateral intact PSBs had more compacted trabecular bone than did control PSBs. Focal repair or remodeling was evident in the palmar aspect of many fractured and contralateral intact PSBs. Fracture coincided with microstructural features and propagated from the flexor to the articular surface.
Conclusions and Clinical Relevance—Fractured PSBs had adapted to high loading but had focal evidence of excessive remodeling and porosity that likely predisposed the horses to complete fracture and catastrophic injury. Detection of focal injury before complete fracture provides an opportunity for prevention of catastrophic injury. Development of diagnostic imaging methods to assess porosity of PSBs may help to identify at-risk horses and allow for modifications of training and racing schedules to reduce the incidence of PSB fracture in Thoroughbred racehorses.
Objective—To investigate the presence or absence of Toll-like receptor (TLR)-2 and TLR-4 in synovial tissues collected from stifle joints (SJs) of dogs with or without osteoarthritis.
Animals—21 purpose-bred research dogs, 3 client-owned dogs with SJ osteoarthritis, and 3 dogs without SJ osteoarthritis.
Procedures—Research dogs underwent arthroscopic surgery in 1 SJ to induce osteoarthritis via cranial cruciate ligament transection (CrCLt; n = 5), femoral condylar articular cartilage groove creation (6), or release of the caudal horn of the medial meniscus (5); 5 dogs underwent sham surgery. Synovial tissue specimens were obtained from both stifle joints of each dog 12 weeks after surgery, and TLR-2 and TLR-4 gene expression were determined via real-time reverse transcription PCR assays. Expression of TLR-4 protein was determined via an immunofluorescence technique in additional specimens obtained from osteoarthritic SJs of dogs with cranial cruciate ligament insufficiency and from dogs with nonosteoarthritic SJs.
Results—Synovial tissues from CrCLt-treated joints had significantly higher TLR-4 gene expression, compared with the contralateral control SJs or any other joint group. TLR-2 gene expression did not differ significantly among groups. Toll-like receptor-4 protein was detected in synovial tissues of osteoarthritic SJs but was rarely evident in nonosteoarthritic SJs.
Conclusions and Clinical Relevance—Increased TLR-4 gene expression in the synovial tissue of SJs with osteoarthritis secondary to CrCLt suggests that activation of innate immunity may play a role in the pathophysiology of SJ osteoarthritis in at least a subset of dogs.
Objective—To determine concentrations of receptor activator of nuclear factor-κB ligand (RANKL) and osteoprotegerin (OPG) in equine chondrocytes and synoviocytes and to quantify changes in the OPG:RANKL ratio in response to exogenous factors.
Sample Population—Samples of articular cartilage and synovium with grossly normal appearance obtained from metacarpophalangeal and metatarsophalangeal joints of 5 adult (1- to 8-year-old) horses.
Procedures—Cell cultures of chondrocytes and synoviocytes were incubated with human recombinant interleukin-1B (hrIL-1β; 10 ng/mL), lipopolysaccharide (LPS; 10 μg/mL), or dexamethasone (100nM) for 48 hours. Negative control cultures received no treatment. Cells and spent media were assayed for RANKL and OPG concentrations by use of western blot and immunocytochemical analyses. Spent media were also assayed for OPG concentration by use of an ELISA.
Results—RANKL and OPG were expressed in equine chondrocytes and synoviocytes in vitro. Cell-associated RANKL and OPG concentrations were not impacted by exogenous factors. Soluble RANKL release into media was significantly increased by hrIL-1β in chondrocyte but not in synoviocyte cultures. Soluble OPG release into media was significantly increased by hrIL-1β and LPS in chondrocyte but not in synoviocyte cultures. The soluble OPG:RANKL ratio was significantly increased by LPS in chondrocyte cultures. Dexamethasone decreased OPG expression in synoviocytes.
Conclusions and Clinical Relevance—RANKL and OPG proteins were expressed in equine articular cells. Release of these proteins may affect osteoclastogenesis within adjacent subchondral bone. Thus, RANKL and OPG may have use as biomarkers and treatment targets in horses with joint disease.
Objective—To evaluate the antioxidant effects of synovial fluid (SF) pooled from metacarpophalangeal joints of healthy horses or horses with various pathological conditions, and to compare them with the antioxidant effects of hyaluronic acid (HA) and chondroitin sulfate (CS).
Sample Population—SF from 1 metacarpophalangeal joint was obtained from 42 horses immediately after humane slaughter. Samples were classified into 3 groups on the basis of origin: healthy joints or joints with chronically damaged cartilage or vascularly congested synovial membranes as detected via macroscopic evaluation.
Procedures—Antioxidant effects were evaluated by use of rat liver microsomal fractions treated with Fe3+-ascorbate as a free radical generator system leading to oxidative stress. Amounts of thiobarbituric-reactive substances and glutathione transferase (GSH-T) conjugation activity were measured.
Results—SF from healthy and chronically damaged joints inhibited microsomal lipid peroxidation, whereas SF from joints with congested synovial membranes had only a slight effect. Hyaluronic acid and CS did not inhibit microsomal lipid peroxidation. Moreover, GSH-T activity was detected in all SF samples, which had similar activity regardless of disease status. All SF samples as well as HA and CS protected rat microsomal GSH-T activity against oxidative damage. Only SF samples from joints with congested synovial membranes protected microsomal thiols against oxidation, an effect also evident with HA and CS.
Conclusions and Clinical Relevance—The antioxidant mechanisms associated with the response to metacarpophalangeal joint damage in horses appeared to act on different targets, depending on whether the damage was acute or chronic.
Objective—To evaluate the ability of signal attenuation–based quantitative magnetic resonance imaging (QMRI) to estimate subchondral bone mineral density (BMD) as assessed via quantitative computed tomography (QCT) in osteoarthritic joints of horses.
Sample Population—20 metacarpophalangeal joints from 10 horse cadavers.
Procedures—Magnetic resonance (MR) images (dorsal and transverse T1-weighted gradient recalled echo [GRE] and dorsal T2*-weighted GRE fast imaging employing steady-state acquisition [T2*-FIESTA]) and transverse single-slice computed tomographic (CT) images of the joints were acquired. Magnetic resonance signal intensity (SI) and CT attenuation were quantified in 6 regions of interest (ROIs) in the subchondral bone of third metacarpal condyles. Separate ROIs were established in the air close to the joint and used to generate corrected ratios and SIs. Computed tomographic attenuation was corrected by use of a calibration phantom to obtain a K2HPO4-equivalent density of bone. Correlations between QMRI performed with different MR imaging sequences and QCT measurements were evaluated. The intraobserver repeatability of ROI measurements was tested for each modality.
Results—Measurement repeatability was excellent for QCT (R 2 = 98.3%) and QMRI (R 2 = 98.8%). Transverse (R 2 = 77%) or dorsal (R 2 = 77%) T1-weighted GRE and QCT BMD measurements were negatively correlated, as were dorsal T2*-FIESTA and QCT (R 2 = 80%) measurements. Decreased bone SI during MR imaging linearly reflected increased BMD.
Conclusions and Clinical Relevance—Results of this ex vivo study suggested that signal attenuation–based QMRI was a reliable, clinically applicable method for indirect estimation of subchondral BMD in osteoarthritic metacarpophalangeal joints of horses.